KR102098843B1 - A fermentation and simulated moving bed process - Google Patents

Abstract

The present invention provides an improved method for the production, separation and recovery of one or more fermentation products from fermentation broth. In addition, the present invention provides a method for increasing the efficiency of the fermentation reaction. In particular, the present invention relates to a fermentation system comprising a simulated moving bed for separation of fermentation products from a fermentation broth, and a corresponding method.

Description

Fermentation and simulating moving bed process {A FERMENTATION AND SIMULATED MOVING BED PROCESS}

Field of invention

The present invention relates generally to systems and methods for producing products, particularly alcohol, by microbial fermentation. In particular, the present invention relates to a fermentation system comprising a simulated moving bed for separation of fermentation products from a fermentation broth, and a corresponding method.

Background of invention

Biofuels for transportation are attractive alternatives to gasoline and are rapidly entering the fuel market as low concentration formulations. Biomaterials derived from natural sources are more environmentally sustainable than those derived from fossil fuel sources (eg gasoline), and their use is so-called fossil carbon dioxide released into the atmosphere as a result of fuel consumption. (CO ₂ ) Reduce the level of gas. In addition, biofuels can be generated locally in many geographic regions and can act to reduce dependence on imported fossil energy resources.

Ethanol has become a major hydrogen-rich liquid transport fuel around the world. Global consumption of ethanol is expected to reach 27.2 billion gallons in 2012, and the global market for the fuel ethanol industry has also been predicted to grow rapidly in the future. This growth is largely due to the growing interest in ethanol in Europe, Japan, the United States and several developing countries.

In the United States, for example, ethanol is used to produce E10, a 10% mixture of ethanol in gasoline. In E10 formulations, the ethanol component acts as an oxygenating agent, improving combustion efficiency and reducing the production of air pollutants. In Brazil, ethanol satisfies about 30% of the transport fuel demand, both in the gasoline blended oxygen supply and in itself pure fuel. In addition, in Europe, environmental concerns surrounding the consequences of Green House Gas (GHG) emissions have led the EU to focus on the consumption of sustainable transport fuels such as ethanol, biomass-derived by EU members. It has been a stimulus to set the goals set out in the law.

Butanediols including 1,2-butanediol, 1,3-butanediol, 1,4-butanediol and 2,3-butanediol can be used as fuel additives for automobiles. They can also be converted relatively easily to a number of other energy products, potentially higher values and / or higher. For example, 2,3-butanediol can be easily converted to 8-carbon dimers that can be used as aviation fuel in a two-step process.

2,3-butanediol induces its versatility from their di-functional backbone, i.e., 2-hydroxyl groups, and is located in the multi-adjacent C-atoms so that the molecule is butadiene , Butydione, acetoin, methyl ethyl ketone, etc., to be converted very easily. These chemical compounds are used as basic molecules to prepare a wide range of industrially produced chemicals.

In addition, 2,3-butanediol can be used as fuel in an internal combustion engine. It is, in some ways, more like gasoline than ethanol. As interest in the creation and application of environmentally sustainable fuels has increased, interest in biological processes to produce 2,3-butanediol (commonly referred to as bio-butanol) has increased.

Most fuel ethanol is the primary carbon source, produced through traditional yeast-based fermentation processes using carbohydrates of crop origin, such as sucrose extracted from sugar cane or starch extracted from grain crops. 2,3-butanediol can also be produced by microbial fermentation of carbohydrate-containing feedstock (Syu MJ, Appl Microbiol Biotechnol 55 : 10-18 (2001), Qin et al., Chinese J Chem Eng 14 (1): 132-136 (2006)). However, the cost of these carbohydrate feedstocks is influenced by their value as human food or animal feed, while cultivation of starch or sucrose-producing crops for ethanol production is not economically sustainable in all regions. Thus, this involves developing technologies that convert lower cost and / or richer carbon resources into biofuel products.

Carbon monoxide (CO) is a major, free-energy-rich by-product of incomplete combustion of organic materials such as coal or products of oil and oil origin. In Australia, for example, the steel industry is reported to generate over 500,000 tons of CO annually and release it into the atmosphere.

It has long been recognized that catalytic processes can be used, which can convert a gas mainly composed of CO and / or CO and hydrogen (H ₂ ) into various fuels and chemicals. However, microorganisms can also convert these gases into fuels and chemicals. This biological process, although generally slower than chemical reactions, has several advantages over catalytic processes, including higher specificity, higher yield, lower energy cost and higher toxicity.

The ability of microorganisms to grow on CO as their sole carbon source was first discovered in 1903. This was later confirmed to be a characteristic of organisms using the acetyl coenzyme A (acetyl CoA) biochemical pathway of autotrophic growth (also known as the Woods-Ljungdahl pathway). Many anaerobic organisms, including carbon monoxide, photosynthesis, methane-producing and acetic acid-producing organisms, contain CO, CO _2, H ₂ , It is known to metabolize to various end products such as methane, n-butanol, acetic acid and ethanol.

Anaerobic bacteria, such as those of the genus Clostridium , have been demonstrated to produce ethanol from CO, CO ₂ and H ₂ via the acetyl CoA biochemical pathway. For example, various strains of Clostridium ljungdahlii that produce ethanol from gases are described in WO 00/68407, EP 117309, US Patents 5,173,429, 5,593,886, and 6,368,819, WO 98/00558 and WO 02/08438 It is described in. The bacterium Clostridium autoethanogenum sp is also known to produce ethanol from gases (Abrini et al, Archives of Microbiology 161, pp 345-351 (1994)).

However, biofuel production by microorganisms by fermentation of gases is always associated with co-generation of acetate and / or acetic acid as a by-product. This acetate / acetic acid has the potential to inhibit the reaction and generally needs to be removed from the fermentation broth. Also, if acetate / acetic acid by-products cannot be used for some other purpose, this can lead to waste disposal problems. Acetate / acetic acid is converted to methane by microorganisms, so it has the potential to contribute to greenhouse gas (GHG) emissions.

Fermentation of gaseous substrates that produce products such as ethanol and 2,3-butanediol is typically performed in a bioreactor containing liquid fermentation broth. The broth contains both microorganisms and nutrients for their growth. Over time, nutrients (including the gas base, itself) are converted to the desired products, but unwanted by-products and cellular debris that can be toxic to microorganisms are produced. Both desired and undesired products can inhibit fermentation efficacy, especially when present at high concentrations.

To reduce the reaction inefficiency caused by inhibiting the fermentation reaction and recover the desired products, the broth is removed from the bioreactor in a continuous or batch process and replaced with fresh nutrient medium. The desired products are typically extracted from the broth using standard extraction methods such as fractional distillation and extractive fermentation. However, these known methods for extracting organic metabolites from fermentation solutions have a number of problems.

Solvent extraction systems, when applied to a weak organic broth, often show poor distribution ratios, which makes separation difficult. Salt saturation can improve the distribution ratio, but complicates the extraction process by requiring the removal of salt from waste aqueous and reduces the cost of consumption if the salt cannot be recovered for reuse. Significantly increase. Liquid pressure membranes (reverse osmosis and nanofiltration membranes) do not show sufficiently high rejection for short chain alcohols, diols, and organic acids. Neither hydrophobic or hydrophilic membranes, either of which can be made tight enough for molecular weight cut-offs to show a clear separation, and both membrane types are strict pre-in fermentation broths. It shows severe particulate fouling in the fermentation broth, which requires filtration.

Distillation is currently the main method for continuous, high purity organics recovery. However, distillation is limited to being used with organic products that have a lower boiling point than water and lack of unfavorable azeotropes. Separation of 2,3-butanediol from aqueous solution by distillation is expensive and difficult due to the high boiling point of 2,3-butanediol (180 to 184 ° C) and high affinity of water. Distillation of ethanol from the fermentation broth yields an azeotrope of ethanol and water (ie, 95% ethanol and 5% water) that cannot be resolved by distillation and requires additional steps and techniques for efficient separation. .

Acetic acid is typically removed by filtering the broth to remove suspended organic material, then passing the broth through an activated carbon column to adsorb acetate. The process requires that the pH of the fermentation broth is reduced to less than about 3 before being transferred through the activated carbon column, converting most of the acetate to acetic acid form. This method of removal is undesirable, as it requires additional steps and addition of pH modifying chemicals to the broth.

Known methods of product recovery are not suitable for recovering major classes of organic products that can be produced through fermentation systems comprising 2,3-butanediol and acetic acid (or cost and / or energy consumption and / or recovery of recovered product). Inefficient in terms of proportions). Thus, recovery is a bottle-neck for commercially viable production of biofuels using microbial fermentation, and new techniques are needed to improve recovery in a more efficient and cost-effective manner.

It is an object of the present invention to provide a process and fermentation system that overcomes or ameliorates at least one of the disadvantages of the prior art, or at least provides it to the public as a useful option.

Summary of the invention

The present invention relates to a method for improving the separation efficiency of one or more fermentation products from a fermentation broth. The present invention provides a method for separating one or more fermentation products from a fermentation broth, where the energy requirement for separation is substantially reduced compared to known methods.

The present invention provides an improved method of separating one or more fermentation products from a fermentation broth by providing an improved system for removing water from a fermentation stream comprising one or more fermentation products.

In a first aspect, the present invention provides a method for separating one or more fermentation products from a fermentation broth, the method comprising

a) fermenting a gaseous substrate in a bioreactor containing a culture of one or more microorganisms to produce a fermentation broth comprising one or more fermentation products;

b) delivering the fermentation broth through a treatment zone run at conditions to produce a treated broth stream, the treated stream being substantially free of biomass;

c) providing at least a portion of the treated broth stream to a simulated moving bed (SMB) module comprising an adsorbent;

d) adsorbing the at least one fermentation product onto the adsorbent and calculating a raffinate containing unadsorbed components of the broth; And

e) desorbing one or more products from the adsorbent to produce a product stream.

In one embodiment of the first aspect, the treatment region comprises at least one heat treatment region. In one embodiment of the first aspect, the treatment region includes a heat treatment region and a filtration region. In one embodiment, the treatment zone removes at least a portion of the suspended and / or soluble biomass from the fermentation broth. In one embodiment, the treatment zone removes substantially all biomass suspended and / or soluble from the fermentation broth. In certain aspects of the invention, the treated broth stream is substantially free of biomass. In certain embodiments, the treated broth stream contains trace biomass.

In one embodiment of the first aspect, the method includes recycling raffinate to the bioreactor.

In a further embodiment of the first aspect, the products are desorbed in step (e) by flushing the adsorbent with a solvent to yield a product-solvent solution. Preferably, the product-solvent solution is substantially free of salts from the fermentation broth. Preferably, the concentration of water in the product-solvent solution is less than 5% by volume, or less than 3% by volume, or less than 1% by volume. In one embodiment, the product-solvent solution is substantially free of water.

Preferably, the products include acids and / or alcohols. In certain embodiments, the solvent is alcohol. According to one embodiment, the product is selected from the group comprising ethanol, acetic acid, 2,3-butanediol, butanol, iso-propanol and acetone. In one aspect, the solvent is selected from the group comprising ethanol, methanol, propanol and methyl t-butyl ether. In a preferred embodiment, the one or more products are selected from the group comprising ethanol, 2,3-butanediol and acetic acid, and the solvent is ethanol.

In one embodiment, the solvent used in the adsorption step is a product of a pre-extracted fermentation process. It will be appreciated that desorption with the product of the fermentation process means that the product is produced without adding additional required separation steps, although additional separation may be required to separate different products of fermentation from the other, One or more products are produced here.

In a particular embodiment, the gaseous substrate is fermented in a bioreactor in step (a) to produce a fermentation broth comprising ethanol and 2,3-butanediol. The fermentation broth is delivered to the treatment area, where at least a portion of the biomass and / or soluble protein is removed from the fermentation broth to provide a treated stream. In a particular embodiment, the treated stream is flowed to SMB, where at least some of the ethanol and 2,3-butanediol is absorbed from the treated stream to produce a raffinate stream. The solvent is delivered through an adsorber to desorb the ethanol and 2,3-butanediol to provide an extraction stream. In a further embodiment, the extraction stream is delivered through a recovery zone operated under conditions to provide an ethanol stream and a 2,3-butanediol stream. In particular embodiments, at least a portion of the raffinate stream is passed back to the bioreactor.

In a second aspect, the present invention provides a method for producing and recovering one or more fermentation products from a fermentation broth, the method comprising:

a) fermenting a gaseous substrate in a bioreactor containing a culture of one or more microorganisms to produce a fermentation broth comprising one or more fermentation products;

b) delivering the fermentation broth to a treatment zone operated under conditions for producing a treated broth stream, wherein the treated broth stream is substantially free of biomass;

c) providing at least a portion of the treated broth stream to a simulated moving bed (SMB) module comprising an adsorbent;

d) adsorbing the at least one fermentation product onto the adsorbent to yield a raffinate containing the unadsorbed components of the broth;

e) desorbing the one or more products from the adsorbent to yield a product stream; And

f) recycling at least a portion of the raffinate to the bioreactor.

In one embodiment of the invention, the treatment region of step (b) removes at least a portion of the biomass and / or soluble protein from the fermentation broth to provide a treated broth stream that is substantially free of biomass. In one embodiment, at least a portion of the biomass and / or soluble protein is returned to the bioreactor.

In one embodiment of the present invention, at least a portion of the fermentation broth is passed through a filtration step while exiting the bioreactor, thereby producing a permeate stream. In certain implementations, the permeate stream and processed broth stream are combined before being delivered to the SMB module.

In one embodiment, the raffinate returns to the bioreactor to form part of the liquid nutrient medium. In certain embodiments, the raffinate undergoes a media preparation step before returning to the bioreactor. In certain embodiments, the step of preparing the medium comprises adding one or more nutrients to the raffinate stream.

In certain embodiments, the raffinate is substantially free of products. In preferred embodiments, the raffinate comprises at least 80% H ₂ O, or at least 85% H ₂ O, or at least 90% H ₂ O, or at least 95% H ₂ O. In certain embodiments, the raffinate comprises a trace amount of solvent used to desorb the one or more products from the adsorbent.

In a third aspect, a method of producing and recovering one or more acids is provided, the method comprising:

a) flowing a gaseous material from the liquid nutrient broth into a culture or bioreactor containing one or more microorganisms;

b) fermenting the gaseous substrate to produce a fermentation broth comprising one or more acid (s);

c) delivering the fermentation broth to a treatment region, wherein at least a portion of the biomass and / or soluble protein is removed from the fermentation broth to provide a treated broth stream;

d) flowing the treated broth stream to a simulated moving bed module comprising an adsorbent;

e) adsorbing at least a portion of one or more acids from the treated broth stream to an adsorbent to produce a raffinate stream;

f) desorbing the at least one acid by delivering a solvent through an adsorbent; And

g) delivering at least a portion of the raffinate stream to the bioreactor.

In one embodiment of the third aspect, the adsorbed acid is lactic acid and / or acetic acid, and removal of the acid prevents inhibition and / or collapse of the broth culture. In a particular embodiment, the adsorbed lactic acid and / or acetic acid is desorbed from the absorbent and exits the SMB through an extraction stream. Thus, the pH of the broth is adjusted through the extraction stream, through removal of the lactic acid and / or acetic acid. In one embodiment, removal of the acid from the bioreactor prevents inhibition of the culture of one or more microorganisms.

In one embodiment, one or more acid (s) is desorbed in step (f) with a solvent. According to one embodiment, the solvents used for desorption are ethanol, methanol, propanol and methyl t-butyl ether. In a further embodiment, the solvent used for desorption is a solvent produced by a pre-extracted fermentation process.

In an alternative embodiment of the third aspect, at least a portion of the acid in the treated broth stream is converted to its corresponding salt before being provided to the SMB module. In one embodiment, the acid of the treated broth stream is acetic acid which is converted to sodium acetate using sodium hydroxide. The converted sodium acetate is provided to the SMB module along with the treated broth stream, where the sodium acetate exits the SMB module together with the raffinate and is recycled back to the bioreactor.

In one embodiment, the biomass and / or soluble protein removed from the fermentation broth in step (c) is recycled to the bioreactor.

In one embodiment of the third aspect, one or more acids are removed by the process, whereby the pH of the bioreactor is maintained within a desired range. Microbial growth and metabolite production can be optimized by maintaining the pH in the bioreactor within a desired range. In a particular embodiment, the preferred range is ± 0.5 units of the optimum operating pH. Typically, in acetic acid fermentation, the pH is 6 to 8, or 6.5 to 7.5, or 6.7 to 7.4, or 6.8 to 7.3, or 6.9 to 7.1, or substantially 7.0. According to the first and second aspects of the invention upon fermentation, the pH is between 4.5 and 6; Alternatively 4.61 to 5.9; Alternatively 4.7 to 5.8; Or 4.8 to 5.5. In one embodiment, the pH is substantially maintained at pH 4.8, or pH 5.0, or pH 5.5.

In certain embodiments of the third aspect, the main fermentation product is acetic acid. In certain embodiments, the gas base provided to the reactor is selected from the group consisting of CO, CO, and H ₂ , CO _2, and H ₂ , CO ₂ , CO and H ₂ , or mixtures thereof. In one embodiment of the third aspect, at least one microorganism acetonitrile tumefaciens woodiyi (Acetobacterium woodii), Clostridium Gaetano genum (Clostridium autoethanogenum) Otto, Clostridium buckling unlike (Clostridium ljungdahlii), Clostridium ln seudal Ray ( Clostridium ragsdalei ), Clostridium coskatii , or mixtures thereof.

Step (a) of the third aspect can produce other products such as alcohol. In a particular embodiment, one or more additional fermentation products are adsorbed in step (e). According to one embodiment, the additional product is selected from the group comprising ethanol, 2,3-butanediol, butanol, and iso-propanol.

In any particular embodiment of the above aspects, the treatment step includes at least a filtration step, where at least a portion of the suspended and / or soluble biomass is removed from the fermentation broth before it is delivered to the SMB module. Filtration results in the removal of suspended and / or soluble biomass from the fermentation broth. In certain embodiments, filtration results in a treated broth stream that is substantially free of biomass. Filtration may consist of a route that delivers the broth through a membrane. In one embodiment, flocculation can be induced by adding a flocculant prior to filtration.

In certain implementations, the processing step further comprises at least one heat processing step. It will be appreciated by those skilled in the art that other methods for the removal of biomass from the broth stream can also be used in the treatment step.

Performing the method of the first aspect may result in performing the method of the second aspect, and vice versa.

In a further aspect, the present invention provides a fermentation system, the system comprising at least

a) a bioreactor containing a fermentation broth containing a culture of one or more microorganisms capable of producing one or more fermentation products from a gaseous substrate;

b) a simulated moving bed (SMB) module adapted to be provided with a portion of the fermentation broth;

c) an adsorbent in the SMB module adapted to adsorb one or more fermentation products from a portion of the fermentation broth.

In one embodiment, the system further comprises a treatment module adapted to remove suspended and / or soluble biomass from a portion of the fermentation broth before the broth is received by the SMB module. The processing module includes at least one filtration module. In certain implementations, the treatment module includes a heat treatment module and a filtration module. As indicated above, the SMB module may be provided in or as part of a bioreactor or separately, but may also be in fluid communication with it to accommodate a portion of the broth.

In one embodiment, the system further comprises means for delivering the removed biomass / soluble protein back to the bioreactor. In one embodiment, the system includes means for delivering the raffinate stream exiting the SMB module back to the bioreactor.

In particular embodiments of the third aspect, the bioreactor is configured for gas-based fermentation to produce products comprising acid (s) and / or alcohol (s). In a particular embodiment, the gaseous substrate comprises CO, and optionally H ₂ . In an alternative embodiment, the gaseous substrate comprises CO ₂ and H ₂ .

In particular embodiments of the third aspect, the system comprises an adjusting means and a processing means, wherein parameters including the medium feed rate, the liquid retention time and the substrate feed rate are according to methods known in the art, such as the present invention. Can be controlled, such as the method described in WO2010 / 093262, which is fully incorporated by reference and herein by reference.

In any particular embodiment of the above aspects, the method further comprises treatment of the fermentation broth each removed from the bioreactor or raffinate before or after product removal in the SMB module. In particular embodiments, the treatment may consist of additional components or nutrients (such as vitamin B) added to the raffinate to supplement the nutrient medium prior to recovery to the bioreactor. In addition, the pH of the raffinate can be adjusted prior to recovery to the bioreactor and adjusting the pH of the broth in the bioreactor.

In any particular embodiment of the above aspects, the adsorbent is a fluorinated carbon adsorbent. In an alternative embodiment, the adsorbent is an activated carbon adsorbent. In other embodiments, the adsorbent is a C18 surface modified silica gel.

The adsorbents mentioned above are examples of suitable adsorbents and are not intended to be an exhaustive list. The skilled artisan will appreciate that any adsorbent material with suitable selectivity and hydrophobicity for use in the SMB process defined herein can be used.

Although it is believed that it is desirable for SMB to be in fluid communication with it while being separated from the bioreactor, the SMB can be provided in the bioreactor. When SMB is included in the bioreactor, preferably, the SMB is suspended in the broth and / or kept separate from soluble biomass. For example, a portion of the broth can be separated from the rest by a membrane so that the SMB communicates with the fermentation products but not with suspended or soluble biomass that can affect the performance of the SMB. Additionally or alternatively, the feed of the SMB can be provided with the filter to the same end. This applies to all aspects of the invention.

Surprisingly, the SMB process is advantageous for separating the desired product from the treated broth stream and / or fermentation broth comprising a diluted concentration of organic product. In one embodiment of the present invention, the concentration of ethanol and / or 2,3-butanediol in the fermentation broth and treated broth stream is 30% by weight or less in water, or less than 15% by weight in water. In one embodiment, the fermentation broth or treated stream contains 2-10% by weight of ethanol / 2,3-butanediol in water, wherein ethanol to 2,3-butanediol is in a ratio of 5; 1 to 1; 1 exist. In a preferred embodiment, the fermentation broth or treated stream contains 6% by weight of ethanol / 2,3-butanediol in water, wherein ethanol to 2,3-butanediol is present in a 1: 1 ratio. It has also been surprisingly found that 2,3-butanediol concentrations of less than 2% by weight can be separated using the SMB process. In particular embodiments, the adsorbent adsorbs at least about 50%, about 60%, about 70%, about 80%, about 90%, about 95%, about 99% or substantially 100% of the fermentation product from the broth. In one embodiment, the adsorbent adsorbs 50 to 100%, or 60 to 95%, or 70 to 90%, or 70 to 100% of the fermentation product from the broth.

In particular embodiments of the present invention, the adsorbent preferably adsorbs at least 6.0 W / W, or at least 7.0 W / W, or at least 8.0 W / W, or at least 9.0 W / W, or at least 10.0 W / W of ethanol. Will have a ratio. In certain embodiments, the adsorbent has an ethanol adsorption ratio of 6.0 to 10.0 W / W, or 7.0 to 10.0 W / W, or 6.0 to 9.0 W / W, or 7.0 to 9.0 W / W.

According to the invention, the adsorbent is preferably at least 9.0 W / W, or at least 10.0 W / W, or at least 12.0 W / W, or at least 16.0 W / W, or at least 18.0 W / W, or at least 20.0 W / W It will have a 2,3-butanediol adsorption ratio. In certain embodiments, the adsorbent is 9.0 to 20.0 W / W, or 12.0 to 20.0 W / W, or 10.0 to 18.0 W / W, or 12.0 to 18.0 W / W, or 16.0 to 20.0 W / W, 2,3 -Has an adsorption ratio of butanediol.

In a particular embodiment of the invention, the temperature at which organic products are adsorbed on the adsorbent is 20 ° C to 75 ° C, or 25 ° C to 40 ° C, or 25 ° C to 35 ° C. In a preferred embodiment, the temperature at which organic products are adsorbed to the adsorbent is about 25 ° C. As will be appreciated, it is significantly lower than required to separate the product by distillation.

In certain embodiments of the invention, the temperature at which the product is desorbed from the adsorbent is 20 ° C to 120 ° C, or 20 ° C to 110 ° C, or 25 ° C to 100 ° C, or 40 ° C to 100 ° C, or 40 ° C to 90 ° C Less than. In certain embodiments, the temperature at which the product desorbs from the adsorbent is about 90 ° C.

In a particular embodiment of the invention, the pressure at which the product is adsorbed on the adsorbent is less than 200 psig (1,379 kPag), or less than 150 psig (1,034 kPag) or less than about 100 psig (689 kPag), or about 50 psig (345 kPag) Less than. In certain embodiments, the pressure at which the product is adsorbed on the adsorbent is 14.7 to 200 psig (101 to 1,379 kPag). Embodiments of the present invention find special application in the separation of organic products of gaseous fermentation, such as acids, alcohols and diols, generally from aqueous fermentation broths. In particular acetic acid, ethanol and 2,3-butanediol are produced by gas-based fermentation involving CO and can be separated from aqueous organic streams using the present invention.

In further embodiments of the above aspects, an alcohol product such as ethanol is extracted from a portion of the broth removed from the bioreactor (or other portion of the broth) prior to delivery of the broth to the SMB module, and optionally to the filtration module. Preferably, ethanol is extracted from the broth by distillation. In a particular embodiment, the extracted alcohol is used as a desorbent in the SMB module.

In further embodiments of the above aspects, the SMB module is regenerated after adsorption of products and / or acids. In a particular embodiment, the adsorbent removes substantially all desorbent. In particular embodiments, the adsorbent removes the desorbent by steam stripping. Steam stripping may occur prior to adsorption to produce a condensed stripping solution removed from the system, or it may occur with an adsorption step in which the condensed stripping solution is carried out of the system along with the raffinate. The condensed stripping solution or desorbent-containing raffinate is distilled to recover the extracted desorbent, which is returned to the process.

The gaseous substrate can include the gas obtained as a by-product of an industrial process. In certain embodiments, the industrial process includes ferrous metal product manufacturing, non-ferrous product manufacturing, petroleum refining processes, gasification of biomass, gasification of coal, electrical power generation, carbon black generation, ammonia generation, It is selected from the group consisting of methanol production and coke production. In one embodiment of the invention, the gas base is a synthetic gas. In one embodiment, the gas substrate comprises a gas calculated from a steel mill.

In a particular embodiment, the gaseous substrate is a gaseous substrate containing CO. In further embodiments, the substrate is at least about 15% by volume CO to 100% by volume CO, e.g., 20% by volume CO to 100% by volume CO, e.g., 43% by volume CO to 95% by volume CO, e.g. For example, 75 volume% CO to 95 volume% CO, or, for example, 80 volume% to 90 volume% CO. In one particular embodiment, the gaseous substrate comprises about 95% CO. If the substrate also contains CO ₂ and H ₂ , lower CO levels, such as 6%, are contemplated. In other embodiments, the substrate stream comprises a 2% to 13% H ₂ concentration.

In various embodiments, fermentation is performed using a microbial culture comprising one or more strains of carbon monoxide nutrition bacteria. In various embodiments, the carbon monoxide nutritional bakteriaheun Clostridium (Clostridium), no Pasteurella (Moorella), oxo bakteo (Oxobacter), pepto streptococcus (Peptostreptococcus), acetonitrile tumefaciens (Acetobacterium), oil cake Te Solarium (Eubacterium), Or Butyribacterium . In one embodiment, the carbon monoxide nutrition bacterium is Clostridium autoethanogenum . In a particular embodiment, the bacteria have the identification properties of accession number DSMZ10061 or DSMZ23693.

The present invention is also directed to any or all combinations of two or more of the parts, elements or contents, individually or collectively, the parts, elements and contents mentioned or indicated in the present application specification, and the present invention. Included where specific integers having equivalents known in the art related to the invention are mentioned, and such known equivalents are intended to be included individually as set forth herein.

These and other aspects of the invention should be considered in all of these novel aspects, and it will be apparent that it is given by way of example only with reference to the accompanying drawings, which are:
1 is a schematic diagram of a fermentation system according to an embodiment of the present invention;
2 includes a schematic diagram of a fermentation system according to an embodiment of the present invention, wherein the SMB module is connected to gas fermentation to extract fermentation products such as ethanol and 2,3-butanediol.

Detailed description of the invention

Definitions

Unless otherwise indicated, the following terms are defined throughout the specification as follows:

Raffinate-the remainder of the fermentation broth after the fermentation products have been adsorbed to the adsorbent.

Fermentation broth or broth-a mixture of ingredients (including broth culture and nutrient medium) found in the bioreactor.

Nutrient medium-solution added to fermentation broth containing nutrients and other ingredients suitable for the growth of microbial cultures.

Broth Culture-Microbial culture present in fermentation broth.

Broth culture density-The density of microbial cells in the fermentation broth.

Gas substrates comprising carbon monoxide-and similar terms include any gas containing carbon monoxide. The gaseous substrate will typically contain a significant proportion of CO, preferably at least about 5% by volume to about 100% by volume CO.

Acid-As used herein, this term includes the carboxylic acid form. Acetic acid in the form of acetate is not suitable for use with the adsorbent process of the present invention. Acetate present in the fermentation broth can be converted to the acid form by pH adjustment. The molecular ratio of acetic acid to acetate in the fermentation broth depends on the pH of the system.

Bioreactor or fermenter-is a continuous stirred tank reactor (CSTR), immobilized cell reactor (ICR), trickle bed reactor (TBR), moving bed biofilm reactor (Moving Bed Biofilm) For membrane reactors such as Reactor: MBBR, Bubble Column, Gas Lift Fermenter, Hollow Fiber Membrane Bioreactor (HFMBR), static mixer, or other vessel or gas-liquid contact Fermentation devices consisting of one or more containers and / or towers or piping arrangements, including other suitable devices.

Second or Second Bioreactor-As used herein, these terms are intended to encompass any number of additional bioreactors that may be connected in series or in parallel with the first and / or second bioreactor. Is intended

Fermentation, fermentation process or fermentation reaction-and similar terms, as used herein, are intended to include both the growth phase of the process and the product biosynthesis phase. In some embodiments, the bioreactor can include a first growth reactor and a second fermentation reactor. As such, the treatment, or addition, of components to the fermentation reaction should be intended to relate to each or both of these reactors.

Distribution Ratio-As used herein, it is intended to define the ratio of the concentration of a substance of A in its single form in the extract to its concentration ratio in the same form in different phases in equilibrium, which is Appears as:

. .Ⅰ.

. .

(IUPAC.Compendium of Chemical Terminology, 2 ^nd ed. (The "Gold Book" .Compiled by AD McNaught and A, Wilkinson, Blackwell Scientific Publications, Oxford (1997) .XML on-line corrected version: hip: // goldbook. iupac.org (2006) created by M.Nic, J. Jirat, B.Kosta; updates compiled by A.Jenkins.ISBN 0-9678550-9-8)

Components of Nutritional Medium-As used herein, it is intended to refer to any substance provided in a liquid nutritional medium, which assists the growth of microorganisms and includes, but is not limited to, vitamins, trace metals and minerals. Includes. Aqueous organic stream, as used herein, is intended to define an aqueous stream comprising one or more organic products of a fermentation process. Examples of organic products include but are not limited to ethanol; 2,3-butanediol; Acetic acid; Propanol; Butanol; Isopropanol and acetone, compounds that are highly compatible with water, ie to a high degree of solubility in water.

Throughout the specification and any claims, unless the context requires otherwise, the words "comprises", "comprising", and similar words, that is, the exclusive meaning of "including but not limited to" And should be interpreted in a comprehensive sense.

The present inventors have confirmed that SMB can be advantageously applied to extracting organic compounds such as alcohols, diols, and organic acids having high affinity for water, generally from aqueous solutions, and developed a process for this. . So far, SMB has only been used to separate organic compounds from organic solvents or to extract organic compounds from aqueous solutions, where the organic component has a low affinity for water. Alcohols, diols, and organic acids have low carbon chain length and high polarity, so these chemicals tend to have high affinity for water, i.e., are substantially completely soluble in water. . In addition, these compounds are typically produced in low concentration solutions containing impurities (ie, 10% w / w). Fermentation solutions often contain various inorganic compounds as well as suspended and soluble biomass contaminants that limit adsorption by physically blocking or competing for the adsorbent surface. At least some preferred embodiments of the invention aim to overcome at least one of these limitations by providing a process and system having at least one of optimized adsorbent selectivity, capacity, mass transfer ratio, and long-term stability. The invention also advantageously provides an SMB module that is optimized for continuous operation, thereby reducing SMB capital consumption and operating costs.

This method of coupling gas fermentation to SMB technology for extracting fermentation products offers several advantages over known separation methods.

Regenerated, continuous adsorption reduces the amount of adsorbent and solvent / desorbent and energy consumption. Running costs are significantly lower than conventional unit processes such as evaporation, solvent extraction and crystallization.

Compared to fixed beds, SMB has a much more effective volume of functional adsorbent. In a batch (fixed layer) process, a liquid composition at a given bed level periodically changes over time, and most of the layer is not active at a given time. During a continuous process using SMB extraction, the composition at a given level is fixed, and the entire layer performs useful functions.

Fermentation products can be desorbed from the adsorbent by delivering a desorbent / solvent on the adsorbent to yield a product-solvent solution. The present invention is conventional in that it separates organic products into high yields and purity, with minimal to no carry-over of solvents (e.g., water from broth) and / or unwanted solutes. It has additional advantages over separation technology.

An additional advantage of SMB is its ability to simultaneously extract one or more products from solution. Optimization of the adsorbent layer allows SMB to clearly extract multiple organic products under the same process conditions that cannot be performed using existing extraction methods.

In broad terms, the present invention provides a method of separating one or more fermentation products from a fermentation broth using a simulated moving bed (SMB) module comprising an adsorbent.

Embodiments of the present invention find special application in the separation of aqueous organic products of gaseous fermentation such as acids, alcohols and diols from fermentation broth. In particular, acetic acid, ethanol and 2,3-butanediol are produced by gas-based fermentation comprising CO, which can be separated from the aqueous organic stream using the present invention.

Known solvent extraction systems show poor partition coefficients when using broths with low organic product concentrations. Salt saturation can improve the distribution efficiency, but this adds to the cost of removing salt from aqueous waste, which significantly increases the cost of consumption if the salt cannot be recovered for reuse. SMB simplifies the extraction process by requiring minimal chemical consumption and broth treatment due to selective adsorption / desorption of fermentation products. In a particular embodiment of the present invention, if the desorbent chemical used is an organic product from fermentation, the SMB process substantially does not require chemical consumption.

In particular embodiments of the present invention, the methods include the step of filtering the fermentation broth prior to delivery of the fermentation broth to the SMB module. Filtration results in removal of suspended and / or soluble biomass from the fermentation broth. Filtration can pass broth through membranes including, but not limited to, nano filtration and ultra filtration membranes, or may be bypassed by fermentation broth denaturation, or other filtration methods known in the art. Aggregation can be induced by adding a flocculant prior to filtration. Filtration can be performed as a separate module or included as part of the SMB module.

Examples of suitable adsorbents are fluorinated carbon adsorbents. In certain embodiments, the adsorbent is an activated carbon adsorbent or a fluorinated carbon adsorbent. In other embodiments, the adsorbent is a C18 surface modified silica gel.

Preferably, the adsorbent can be any adsorbent material capable of separating water from the denatured fermentation broth. Suitable adsorbents include fluorinated carbon adsorbents. Examples of suitable fluorinated carbon adsorbents include surface fluorinated carbon adsorbents, such as ORSCNCB4FL5GR and ORSNCB4FLGR (available from Orochem Technologies, Inc.), hereinafter referred to as FC-5 and FC-1 respectively. Other suitable adsorbents include activated carbon adsorbents. Examples of activated carbon adsorbents include ORSNCB4GR (available from Orochem Technologies, Inc. of Lombard, Ill.) And is hereinafter referred to as E-325. The C18 surface modified silica gel also has properties suitable for use in the SMB process described herein. An exemplary C18 surface modified silica gel is RELIASIL 5 micron C18 (available from Infochroma, Zug, Switzerland).

By using optimized adsorbents and conditions, the inventors have shown that the fermentation product can be extracted from the broth in a high yield and efficient way compared to conventional extraction techniques. Optimized hydrophobic adsorbents have been found by the inventors to show high capacity and selectivity for organic compounds such as ethanol, propanol, butanol, acetic acid, 2,3-butanediol and acetone. The method also results in substantially complete rejection of water and inorganic salts present in solution.

In particular embodiments, the adsorbent adsorbs fermentation products from at least about 50%, about 60%, about 70%, about 80%, about 90%, about 95%, about 99% or substantially 100% broth.

The products can be separated from the product mixture desorbed by standard methods known to those skilled in the art, such as distillation. For example, ethanol has a boiling point of 78.8 ° C and acetic acid has a boiling point of 107 ° C. As a result, ethanol and acetic acid can be easily separated from each other using a volatile method such as evaporation. Acetate can be recovered by adsorbing onto activated carbon. Another example of separation is an organic-solvent nanofiltration membrane. They enable size separation of two solvent components from each other through pressure filtration.

SMB requires solvent desorbents to remove organic products from adsorbents. Suitable desorbents enable near complete desorbent regeneration while ensuring clear separation from the adsorbent and then from the desorbed product with only slight changes to process conditions. Preferably, the desorbent is selected from the group comprising methanol, ethanol, propanol, and methyl t-butyl ether. In preferred embodiments, the desorbent is methanol or ethanol.

In a particular embodiment, the solvent used for desorption is a solvent produced by the fermentation process and previously extracted. This reduces consumable costs and possible waste disposal needs. Using the extracted product as a solvent reduces the likelihood of unwanted solvent / desorbent or solvent / desorbent contaminants being recycled to the broth which can inhibit fermentation efficiency. In a particular embodiment, the solvent is ethanol produced by fermentation or by a linked fermentation process. These solvents can be extracted from the removed site of the broth prior to delivery to the SMB module or can be obtained from other sites of the broth.

Since SMB relies on the molecular interaction between the target product and the adsorbent surface, its separation performance, unlike distillation, does not require high temperatures. SMB enables continuous recovery of organic compounds from broth hydrocarbons and aqueous solutions without significantly requiring heat or pressure. This can reduce energy consumption and can also result in a reduction in greenhouse gas emissions. Low temperature and pressure requirements also enable recirculation of raffinate with minimal treatment while avoiding deterioration of broth nutrients.

In a particular embodiment of the invention, the temperature at which the product is adsorbed on the adsorbent is between 25 ° C and 75 ° C. In a preferred embodiment, the temperature at which the organic product is adsorbed to the adsorbent is about 25 ° C. In a further particular embodiment of the invention, the temperature at which the product desorbs from the adsorbent is between 25 ° C and 120 ° C. In certain embodiments, the temperature at which the product desorbs from the adsorbent is about 90 ° C. In a particular embodiment of the invention, the pressure at which the product is adsorbed to the adsorbent is less than 200 psig (1,379 kPag) or less than 150 psig (1,034 kPag) or about 100 psig (689 kPag).

In an embodiment of the invention, the raffinate is recycled to the bioreactor. Before recycling, the raffinate may be treated, and the treatment may consist of additional components or nutrients (eg, vitamin B) added to the raffinate to supplement the nutrient medium. The pH of the raffinate can be adjusted before returning to the bioreactor to adjust the pH of the broth in the bioreactor.

Controlling the pH in the fermentation reactor is an important factor that can affect a number of variables, such as reaction rate and product formed. Although the microorganisms involved in fermentation will often produce products over a wide range of pH, maintaining the optimum pH for special reaction conditions can maximize growth and / or production efficiency. Build-up of acids such as acetic acid and lactic acid can inhibit fermentation and, if not identified, can lead to disruption of microbial culture.

In broad terms, the present invention also provides a method for adjusting the pH of a fermentation broth in a bioreactor using a simulated moving bed (SMB) module, preferably comprising an adsorbent for removing a portion of the broth comprising acid. do.

The method can allow the pH of the fermentation broth to be adjusted continuously without requiring the addition of an oxidizing agent or alkalizing agent, or at least reducing the need for it. This reduces the cost of consumption while reducing the waste treatment that may be required to remove the formulation.

In a further embodiment, the present invention provides a method of adjusting the pH, whereby the degree of adjustment of the pH of the broth is determined by the amount of one or more acids extracted from the removed site of the broth.

In acid-producing fermentations, accumulation of acid in the bioreactor can result in inhibition or disruption of microbial cultures. According to one aspect of the invention, the SMB module can be used as an extended cell recirculation system to strip excess acid from the culture while allowing recovery of essential biomass and soluble proteins. In certain embodiments, substantially all acetic acid produced in the fermentation broth is removed through the process. In certain embodiments, when the pH in the bioreactor rises above a desired range, acetic acid can be returned to the reactor in the form of acetate. In certain embodiments, it is possible to adjust the pH in the bioreactor by controlling the amount of acid stripped from it as the broth stream is delivered through the SMB module. If the pH in the reactor falls below the desired level, more acid is stripped from the fermentation broth to return the pH to the desired range. The pH of the bioreactor depends on the type of fermentation. In acetic acid fermentation, if acetic acid is the main fermentation product, the pH range should be maintained between about pH 6 and about pH 7.5. In alcoholic fermentation, when one or more alcohols are the main fermentation product (first and second aspects of the present invention), the pH is maintained between about pH 4.5 and about pH 5.5. In certain embodiments, the process is a continuous process.

The present invention also provides a fermentation system of the exemplary embodiment schematically shown in FIG. 1. The system comprises a bioreactor 1 containing a fermentation broth 2 containing microorganisms capable of producing one or more fermentation products from a gaseous base 3 which can be fed to the bioreactor 1 through a suitable inlet. Includes.

A portion of the broth 2 is fed to the broth 2 in the bioreactor 1 through a filtration module 4 adapted to remove suspended and / or soluble biomass from the fermentation broth. The removed concentrated biomass can be recycled back to the bioreactor 5. The biomass-depleted broth is delivered to a simulated moving bed module 6 comprising an adsorbent adapted to adsorb one or more fermentation products from the biomass-depleted broth, resulting in a raffinate stream (not adsorbed to the adsorbent) (Filtered biomass-depleted broth). The raffinate is recycled to the bioreactor 1 7. The desorbent 8 desorbs the fermentation product which is transferred on the adsorbent and removed in the concentrated metabolism stream 9 which can be applied in a further separation step. The adsorbent 6 removes all residual desorbent (or "regenerated") by steam stripping 10 prior to subsequent adsorption, or with an adsorption step to perform a system with raffinate. In one embodiment, the pH can be adjusted by removing the fermentation product (eg, acid) from the broth using the system described above and shown in FIG. 1.

Another embodiment of the invention is schematically illustrated in FIG. 2. For ease of reference, reference numbers have been used similarly for similar contents. The system comprises a bioreactor 1 containing a fermentation broth 2 containing microorganisms capable of producing one or more fermentation products from the gaseous substrate 3. A portion of the broth may be delivered directly to a filtration module 4 that is adapted to a) remove at least a portion of the suspended and / or soluble biomass from the fermentation broth. Optionally, volatile products such as ethanol are distilled 15 before being passed to the filtration module 4. The solid biomass is removed from the filtration module 4 and can be treated or recycled to the fermentation broth. The evaporated product can be transferred to SMB module 6 and used as a desorbent.

The biomass-depleted broth is delivered to an SMB module 6 comprising an adsorbent adapted to adsorb one or more fermentation products of the broth. The raffinate is recycled 7 times through the media preparation module 20 to the bioreactor where treatment can be performed to optimize the media feed. It will be appreciated that such a module 20 can be added to the FIG. 1 implementation.

The desorbent 8 is introduced into the SMB module 6 and transferred on the adsorbent to desorb the fermentation products. The desorbent may be a fermentation product 17 or 22 such as ethanol or a consumable 23 such as methanol or water. After desorption, the fermentation products are removed in a concentrated metabolite stream 9 for further separation in separation module 25, providing purified products 26 such as ethanol and 2,3-butanediol, respectively, provided only as examples. And (27). The desorbent is removed 28 together with the waste exhaust gas 29 and the desorbent can be collected 30 and recycled.

The adsorbent is regenerated using steam 10 and heated exhaust gas 32. The heated exhaust gas can be obtained by transferring the exhaust gas 33 from the bioreactor through the heat exchanger 34. The concentrated vapor and desorbent can be transferred to a biomass stripped broth for further processing.

In a particular embodiment, the gaseous substrate is at least about 15% by volume CO to 100% by volume CO, for example 20% by volume CO to 100% by volume CO, for example 43% by volume CO to 95% by volume CO, for example For example, 75 volume% CO to 95 volume% CO, or, for example, 80 volume% to 90 volume% CO. In one particular embodiment, the gaseous substrate comprises about 95% CO. If the substrate also contains CO ₂ and H ₂ , lower CO levels, such as 6%, can be considered. In other embodiments, the substrate stream comprises a concentration of H ₂ from 2% to 13%.

Although the description herein focuses on the production of ethanol, 2,3-butanediol and / or acetic acid using CO as the main substrate, i.e., special embodiments of the invention, it is the production of alternative alcohols or acids or alternative compounds. It should be recognized that it is applicable. It is also contemplated to use alternative substrates, including those that would be known to those skilled in the art related to the present invention. For example, gaseous substrates containing carbon dioxide and hydrogen can be used. Further, the present invention may be applicable to fermentation to produce butyrate, propionate, caproate, ethanol, propanol, and butanol. The method can also be used to produce hydrogen. By way of example, these products are free Pasteurella, Clostridia, Rumi Noko Syracuse, acetonitrile tumefaciens, oil cake Te Solarium, butyric Li tumefaciens, oxo bakteo, metadata of labor LE key (Methanosarcina), and Dessel picture Do Coolum (Desulfotomaculum) It can be produced by fermentation using microorganisms from the genus.

Separation of simulated moving bed

The simulated moving bed (SMB) is a separation technique based on adsorption and desorption of target organic solutes from a solution. The technology was developed in the 1950s to purify industrial chemicals. SMB was adopted due to a 10-fold reduction in energy demand and a 5-fold increase in product throughput compared to fixed bed adsorption. SMB development was accelerated by UOP for the separation of organic components with similar boiling and / or azeotropic properties (eg, Sorbex ^™ and MX ^™ methods). Optimization of adsorbent properties enables SMB to be used to extract organic fermentation products from aqueous fermentation broth solutions, where it has been found that the organic component has a high affinity for water.

The SMB runs continuously by circulating and recirculating the continuous stream of broth through the bed by the use of multi-port valve or rotary valve fluid control, while fixing the two or more columns containing the adsorbent bed. If elution is not sufficient to extract the desired product (s) at the desired purity, over the sum of successive columns, the stream can be directed to be delivered through the column an additional number of times until the desired extraction is achieved. have. Thus, a fully considered timed switching of the valve to re-direct the broth stream simulates the movement of the adsorption bed. The remainder of the broth (mainly including water and salt) is called raffinate and can be removed or recycled for disposal.

The multi-delivery approach of SMB is useful, for example when the affinity between compounds is high in the separation of chiral agents.

The SMB process consists of three main steps, which are:

Adsorption step-the feed solution is transferred on an adsorbent to adsorb the organic product onto the surface. The resulting raffinate is removed from the system.

Desorption step-the desorbent solvent is transferred onto the adsorbent to extract the organic product from the surface, and the resulting solution is typically removed by distillation for separation.

Regeneration Step-The adsorbent is removed with all residual desorbent, either prior to adsorption via steam stripping, or with the adsorption step if performed with a system with raffinate. The extracted stripping agent is recovered by distilling the concentrated stripping solution or the raffinate containing the stripping agent, which is returned to the above process.

A rectification stage can be provided between the adsorption step and the desorption step. During the rectification step, at least a portion of the product on the adsorbent will move below the surface of the adsorption column and collect at the bottom of the column. This allows the use of lower desorbents in the desorption step.

As will be appreciated by those skilled in the art, the simulated mobile bed module mentioned herein may include a number of different SMB designs. Exemplary SMB module designs that may be suitable for integration into the methods and systems of the present invention are described, for example, in US 3268605, US3706812, US5705061 and US6004518, the entire contents of which are incorporated herein by reference. Additional devices, as would be appreciated by those skilled in the art, may also be incorporated into an SMB module to aid flow distribution (eg, devices described in US6979402) or provide other benefits.

Although a simulated mobile bed system is mentioned herein, the present invention is also intended to cover the use of fermentation coupled with an actual mobile bed system as described in US6979402 B1, which relies on the same mobile bed concept.

Fermentation

Adopting certain embodiments of the present invention uses gas streams produced by one or more industrial processes. These processes include steel fabrication processes, particularly those that produce a gas stream with a high CO content or a CO content above a certain level (e.g., 5%). According to these embodiments, acetogenic bacteria are preferably used to produce acids and / or alcohols, especially ethanol or butanol, in one or more bioreactors. Those skilled in the art will recognize that the present invention can be applied to a variety of industrial or waste gas streams, and also to vehicles equipped with the above-described internal combustion device. In addition, those skilled in the art will immediately recognize the present disclosure and recognize that the present invention can be applied to other fermentation reactions, including those using the same or different microorganisms. Accordingly, the scope of the invention is not limited to the specific embodiments and / or applications described, but is instead understood in a broader sense; For example, it is intended that the source of the gas stream is also not limiting, except that at least its components are available to feed the fermentation reaction. The present invention has particular application in improving the production and / or overall carbon capture of ethanol and other alcohols from gaseous substrates comprising CO. Processes for producing ethanol and other alcohols from gaseous substrates are known. Exemplary processes include, for example, those described in WO2007 / 117157, WO2008 / 115080, WO2009 / 022925, WO2009 / 064200, US 6,340,581, US 6,136,577, US 5,593,886, US 5,807,722 and US 5,822,111, each of which is incorporated herein by reference. Is included.

Many anaerobic bacteria are capable of fermenting CO into alcohols, diols and acids and are known to be suitable for use in the present invention. Examples of such bacteria suitable for use in the present invention are of the genus Clostridium, such as Clostridium ljungdahlii, e.g. WO 00/68407, EP 117309, U.S. Pat.Nos. 5,173,429, 5,593,886, And 6,368,819, WO 98/00558 and WO 02/08438, Clostridium carboxydivorans (Liou et al., International Journal of Systematic and Evolutionary Microbiology 33: pp 2085-2091), Clostri comprises: (pp 345-351 et al Abrini, Archives of Microbiology 161) Stadium ln seudal ray (Clostridium ragsdalei) (WO / 2008 /028055) and Clostridium auto Gaetano genum (Clostridium autoethanogenum) on. Other suitable bacteria are those of the genus Murella , such as Moorella sp HUC22-1 (Sakai et al, Biotechnology Letters 29: pp 1607-1612), and Carboxydothermus Genus Svetlichny, VA, Sokolova, TG et al (1991), Systematic and Applied Microbiology 14: 254-260. Additional examples include Moorella thermoacetica , Moorella thermoautotrophica , Ruminococcus productus , Acetobacterium woodii , Eubacterium limo breath (Eubacterium limosum), butynyl Li tumefaciens methyl trophy glutamicum (Butyribacterium methylotrophicum), oxo bakteo program penny Kii (Oxobacter pfennigii), meta labor LE key bar Kerry (Methanosarcina barkeri), meta labor LE key acetoxy TiVo lance (Methanosarcina acetivorans ), Desulfotomaculum kuznetsovii (Simpa et. Al. Critical Reviews in Biotechnology, 2006 Vol. 26.Pp41-65). It can also be understood that other acetogenic anaerobic bacteria can be applied to the present invention as understood by those skilled in the art. It will also be appreciated that the present invention can be applied to a mixed culture of two or more bacteria.

One exemplary microorganism suitable for use in the present invention is Clostridium autoethanogenum. In one embodiment, Clostridium autoethanogenum is Clostridium autoethanogenum with the identification characteristics of strains deposited with identification deposit number 19630 to the German Resource Center for Biological Material (DSMZ). In another embodiment, Clostridium autoethanogenum has the identifying properties of DSMZ Accession No. DSMZ 10061 or DSMZ 23693. The laboratory strain of this bacterium is known as LZ1561.

In one embodiment, the microorganism is selected from the group of carbon monoxide nutrient acetogenic bacteria. In certain embodiments, the microorganism is Clostridium autoethanogenum , Clostridium ragdalii , Clostridium ragsdalei, Clostridium carboxydiborans , Clostridium drakei , Clostridium scatologenes , Clostridium coskatii , Butyribacterium limosum , Butyribacterium methylotrophicum , Acetobacterium Acetobacterium woodii, Alkalibaculum bacchii , Blautia producta , Eubacterium limosum, Moorella thermoacetica , Moorela thermoacetica Moorella thermautotrophica , Oxobacter pfennigii , and Thermoan aerobacter kibi ( Thermo anaerobacter kiuvi ).

In one particular embodiment, the microorganism is seed. C. autoethanogenum , C. C. ljungdahlii and C. Ethanol-producing, acetone-producing Clostridia , including Rasdalei and related isolates. These include, but are not limited to, the following strains:

Seed. Autoethanogenum JAI-1 ^T (DSM10061) [Abrini J, Naveau H, Nyns EJ: Clostridium autoethanogenum , sp. nov., an anaerobic bacterium that produces ethanol from carbon monoxide. Arch Microbiol 1994, 4: 345-351], c. Autoethanogenum LBS1560 (DSM19630) [Simpson SD, Forster RL, Tran PT, Rowe MJ, Warner IL: Novel bacteria and methods thereof. International Patent 2009, WO / 2009/064200], C. Autoethanogenum LBS1561 (DSM23693), C. Lungdali PETC ^T (DSM13528 = ATCC 55383) [Tanner RS, Miller LM, Yang D: Clostridium ljungdahlii sp. nov., an Acetogenic Species in Clostridial rRNA Homology Group I. Int J Syst Bacteriol 1993, 43: 232-236]. Lung Dali ERI-2 (ATCC 55380) [Gaddy JL: Clostridium stain which produces acetic acid from waste gases. U.S. Patent 1997, 5,593,886], MR. Gung Dali C-01 (ATCC 55988) [Gaddy JL, Clausen EC, Ko CW: Microbial process for the preparation of acetic acid as well as solvent for its extraction from the fermentation broth. U.S. Patent, 2002, 6,368,819], c. Lung Dali O-52 (ATCC 55989) [Gaddy JL, Clausen EC, Ko CW: Microbial process for the preparation of acetic acid as well as solvent for its extraction from the fermentation broth. U.S. Patent, 2002, 6,368,819], c. C. ragsdalei P11 ^T (ATCC BAA-622) [Huhnke RL, Lewis RS, Tanner RS: Isolation and Characterization of novel Clostridial Species. International Patent 2008, WO 2008/028055], related isolates such as "C. coskatii " [Zahn et al -Novel ethanologenic species Clostridium coskatii (US Patent Application No. US20110229947)], or Seed. Mutant strains such as Tyrando-Acevedo O. Production of Bioethanol from Synthesis Gas Using Clostridium ljungdahlii.PhD thesis, North Carolina State University, 2010. These strains form sub-clusters in Clostridium rRNA cluster I, and their 16S rRNA gene similarly has a homogeneity of 99% or more with a low GC content of about 30%. However, DNA-DNA reassociation and DNA fingerprinting experiments show that these strains belong to individual species [Huhnke RL, Lewis RS, Tanner RS: Isolation and Characterization of novel Clostridial Species. International patent 2008, WO 2008/028055].

All species in this population have a similar shape and size (logarithmic growing cells are 0.5 to 0.7 x 3 to 5 im), mesophilic (optimal growth temperature is 30 to 37 ° C) and stringent. It is very anaerobic [Tanner RS, Miller LM, Yang D: Clostridium ljungdahlii sp. nov., an Acetogenic Species in Clostridial rRNA Homology Group I. Int J Syst Bacteriol 1993, 43: 232-236; Abrini J, Naveau H, Nyns EJ: Clostridium autoethanogenum , sp. nov., an anaerobic bacterium that produces ethanol from carbon monoxide. Arch Microbiol 1994, 4: 345-351; Huhnke RL, Lewis RS, Tanner RS: Isolation and Characterization of novel Clostridial Species. International Patent 2008, WO 2008/028055]. Moreover, they are all in the same pH range (pH 4 to 7.5, optimal initial pH is 5.5 to 6), strong autotrophic growth with similar growth rates in the gas phase containing CO, and ethanol and acetic acid as the main fermentation end products, and It shares the same major phylogenetic properties as metabolic profiles similar to small amounts of 2,3-butanediol and lactic acid formed under certain conditions [Tanner RS, Miller LM, Yang D: Clostridium ljungdahlii sp. nov., an Acetogenic Species in Clostridial rRNA Homology Group I. Int J Syst Bacteriol 1993, 43: 232-236; Abrini J, Naveau H, Nyns EJ: Clostridium autoethanogenum , sp. nov., an anaerobic bacterium that produces ethanol from carbon monoxide. Arch Microbiol 1994, 4: 345-351; Huhnke RL, Lewis RS, Tanner RS: Isolation and Characterization of novel Clostridial Species. International Patent 2008, WO 2008/028055]. Indole production was also observed in all three species. However, the species may include various sugars (e.g. rhamnose, arabinose), acids (e.g. gluconate, citrate), amino acids (e.g. arginine, histidine), or other substrates (e.g. , Betaine, butanol). Moreover, some of these species have been found to be nutritional requirements for certain vitamins (eg, thiamin, biotin), while others are not.

Cultivation of bacteria used in the methods of the present invention can be performed using a specific number of processes known in the art for culturing and fermenting substrates using anaerobic bacteria. By way of example, using gas substrates for fermentation, these processes generally described in the following documents can be used: (i) K. T. Klasson, et al. (1991). Bioreactors for synthesis gas fermentations resources. Conservation and Recycling, 5; 145-165; (ii) K. T. Klasson, et al. (1991). Bioreactor design for synthesis gas fermentations. Fuel. 70. 605-614; (iii) K. T. Klasson, et al. (1992). Bioconversion of synthesis gas into liquid or gaseous fuels. Enzyme and Microbial Technology. 14; 602-608; (iv) J. L. Vega, et al. (1989). Study of Gaseous Substrate Fermentation: Carbon Monoxide Conversion to Acetate. 2. Continuous Culture. Biotech. Bioeng. 34. 6. 785-793; (v) J. L. Vega, et al. (1989). Study of gaseous substrate fermentations: Carbon monoxide conversion to acetate. 1. Batch culture. Biotechnology and Bioengineering. 34. 6. 774-784; (vi) J. L. Vega, et al. (1990). Design of Bioreactors for Coal Synthesis Gas Fermentations. Resources, Conservation and Recycling. 3. 149-160; All of these documents are incorporated herein by reference.

Fermentation may include one or more continuously stirred tank reactors (CSTR), fixed cell reactor (s), gas-lift reactor (s), bubble column reactor (s) (BCR), membrane reactors, e.g. For example, it can be carried out in any suitable bioreactor such as a hollow fiber membrane bioreactor (HFMBR) or trickle bed reactor (s): TBR. Further, in some embodiments of the present invention, the bioreactor (s) may include a first growth reactor in which microorganisms are cultured, a second from which the fermentation broth from the growth reactor is supplied and most of the fermentation product is produced. And a fermentation reactor. In particular embodiments, the second bioreactor is different from the first bioreactor.

According to various embodiments of the present invention, the carbon source for the fermentation reaction is a gas base containing CO. The substrate can be a waste gas containing CO produced as a by-product of an industrial process, or from other sources, such as from automobile fumes. In certain embodiments, the industrial process may include iron metal products such as steel mills, non-ferrous product manufacturing, petroleum refining processes, gasification of coal, electrical power generation, carbon black generation, ammonia generation, methanol generation and coke production. Metal products. In these embodiments, the substrate containing CO can be captured from industrial processes before it is released into the atmosphere using any convenient method. Depending on the composition of the substrate containing CO, it may be desirable to treat it before introducing it into fermentation to remove any unwanted impurities such as dust particles. For example, the gaseous substrate can be filtered or cleaned using known methods.

Alternatively, the substrate containing CO can be supplied from the gasification of biomass. The gasification process involves partial combustion of biomass, with a limited supply of air or oxygen. The resulting gas typically contains mainly CO and H ₂ and a minimum volume of CO ₂ , methane, ethylene and ethane. For example, sugars from sugar cane, or starch from corn or grains, biomass byproducts produced during the extraction and processing of food, or non-food biomass waste produced by forestry can be gasified for use in the present invention. It is possible to produce a gas containing CO suitable for.

Substrates containing CO are in most proportions of CO, for example at least about 15% by volume CO to 100% by volume CO, for example 20% by volume CO to 100% by volume CO, for example 43% by volume CO To 95 vol% CO, for example, 75 vol% CO to 95 vol% CO, or 80 vol% to 90 vol% CO. In one particular embodiment, the gaseous substrate comprises about 95% CO. If the substrate also contains CO ₂ and H ₂ , lower CO levels, such as 6%, can be considered. In other embodiments, the substrate stream comprises a 2% to 13% H ₂ concentration.

Although it is not necessary for the substrate to contain any hydrogen, the presence of H ₂ should not be detrimental to product formation according to the methods of the present invention. In particular embodiments, the presence of hydrogen results in improved overall alcohol production efficiency. For example, in particular embodiments, the substrate may include H ₂ : CO in a ratio of about 2: 1, or 1: 1, or 1: 2. In other embodiments, the substrate stream comprises a 2% to 13% H ₂ concentration. In other embodiments, the substrate stream comprises, or substantially comprises, a low concentration of H ₂ , for example, less than 5%, or less than 4%, or less than 3%, or less than 2%, or less than 1%. Does not contain hydrogen. The substrate may also contain some CO ₂ , such as, for example, about 1% by volume to about 80% by volume of CO ₂ , or 1% by volume to about 30% by volume of CO ₂ . In a particular embodiment, the substrate stream contains CO ₂ and minimal or no CO.

Typically, carbon monoxide will be added in the gas phase to the fermentation reaction. However, the method of the present invention is not limited to the addition of the substance in this state. For example, carbon monoxide can be provided as a liquid. For example, the liquid may be saturated with a carbon monoxide-containing gas or such liquid may be added to the bioreactor. This can be achieved using standard methods. As an example, a microbubble dispersion generator (Hensirisak et.al. Scale-up of microbubble dispersion generator for aerobic fermentation; Applied Biochemistry and Biotechnology Volume 101, Number 3 / October, 2002 ) can be used for this purpose. have.

It will be appreciated that in order for bacterial growth and CO-to-product fermentation to occur, it will be necessary to supply a suitable liquid nutrient medium to the bioreactor in addition to the base gas containing CO. The nutrient medium will contain enough vitamins and minerals to allow the growth of the microorganisms used. Anaerobic media suitable for fermentation of ethanol using CO as the sole carbon source are known in the art. For example, suitable media can be found in the aforementioned U.S. Patent Nos. 5,173,429 and 5,593,886 and WO 02/08438, WO2007 / 117157, WO2008 / 115080, WO2009 / 022925, WO2009 / 058028, WO2009 / 064200, WO2009 / 064201 and WO2009 / 113878 It is described.

It may be desirable to perform the fermentation under conditions suitable to produce the desired fermentation (eg, microbial growth and / or ethanol production). Reaction conditions that can be considered include pressure, temperature, gas flow rate, liquid flow rate, medium pH, medium redox potential, aggregation rate (when using a continuous stirred tank reactor), inoculum level, Maximum gas base concentration to ensure that CO in the liquid phase is not limited, and maximum product concentration to avoid product inhibition. Suitable conditions are described in WO02 / 08438, WO07 / 117157, WO08 / 115080 and WO2009 / 022925.

The present invention includes systems or methods using additional control means and processing means such that parameters including media feed rate, liquid retention time and substrate feed rate are described herein and in methods known in the art, such as herein. It is contemplated that it may be adjusted according to the method described in WO2010 / 093262, which is incorporated by reference in its entirety.

Optimal reaction conditions will depend in part on the particular microorganism used. However, in general, it is preferable to perform fermentation at a pressure higher than ambient pressure. Operation at elevated pressures increases the rate of CO transfer from the gas phase to the liquid phase, where it can be performed by microorganisms as a carbon source for ethanol production. This ultimately means that the retention time (defined as the liquid volume in the bioreactor divided by the input gas flow rate) can be reduced if the bioreactor is maintained at a pressure slightly above ambient pressure.

In addition, the provided CO-to-ethanol conversion rate is partly a function of substrate retention time, and finally achieving the desired retention time in turn represents the required volume of the bioreactor, so the use of a pressurized system Can significantly reduce the volume of the bioreactor required, and consequently the capital cost of the fermentation apparatus. According to the embodiment provided in U.S. Patent No. 5,593,886, the reactor volume can be reduced in a linear proportion to the increase in reactor operating pressure, i.e., a bioreactor running at 10 atmospheres is only 1 of the volume operating at 1 atmosphere. Only / 10 is required.

The advantage of performing gas-to-ethanol fermentation at elevated pressure has also been described. For example, WO 02/08438 describes a gas-to-ethanol fermentation performed under pressures of 30 psig and 75 psig, yielding ethanol productivity of 150 g / l / day and 369 g / l / day, respectively. have. However, it has been found that exemplary fermentations performed using similar media and input gas composition at atmospheric pressure produce 10-20 times less ethanol / liter / day.

It is also preferred that the rate of introduction of the gaseous base containing CO is such that the concentration of CO in the liquid phase is not limited. This is because the result of CO-limited conditions can increase this acetic acid production and reduce ethanol production.

Example

Table 1. Medium composition

Table 2. Trace metal solutions

Table 3. Vitamin solutions

Example 1-Fermentation for recovery of fermentation products

The medium was prepared in volumes of 1.5 L and 1.5 ml of resazurin added according to the compositions described in Tables 1-3. The solution was stirred while heating and degassing with N ₂ . ANa ₂ S drip was started at a rate of 0.1 ml / hr and the temperature of the bioreactor was set to 37 ° C. The pH was adjusted to 5.0 with NH ₄ OH and chromium was added to adjust the ORP to -200 mV. The bioreactor was then fed with RMG (43% CO, 20% CO ₂ , 2.5% H ₂ and 33% N ₂ ) at a flow rate of 50 ml / min. The solution was inoculated with 150 ml of actively growing Clostridium autoethanogenum culture. Once the reactor was continuous, cell recycling was also initiated to yield a bacterial dilution rate of 1.38 days ^-1 and a media flow rate of 2.3 days ^-1 . During operation, product concentration was maximized by increasing stirring (rpm) and gas flow (ml / min). Fermentation was run for 8 days. Table 4 shows the concentration of metabolites in the liquid outflow of the bioreactor.

Table 4. Concentrated metabolite spills

Example 2- Recovery of fermentation products from fermentation broth

Pre-treatment of broth stream

Solid biomass / bacteria were removed from the solution using a 0.1 μm ceramic membrane cross flow filter (GE Healthcare Life Sciences Xampler Microfiltration Cartridge type). After filtration, soluble biomass remained in solution, which must be minimized in order for SMB to function clearly.

A 19 ml guard column containing activated carbon was tested for its ability to remove the remaining biomass and soluble proteins from the feed prior to testing in an SMB device. The protein concentration of the solution was measured before and after the column using BCA analysis and the size distribution of the protein before and after the guard column was evaluated using SDS-PAGE analysis. Prior to delivering the guard column, the protein concentration was about 1000 μg / ml and the protein size was found to be less than 200 kDa, 70 kDa, 40 kDa, 30 kDa, and 2 kDa. The guard column was observed to remove 80% of the soluble protein from the solution and the remaining protein was found to be less than 2 kDa in size.

The guard layer was measured to have 4 g of adsorbed protein and other soluble biomass such as DNA and enzymes. Protein was desorbed from the adsorbent layer using 5 column volumes of methanol and about 3.7 g of protein was removed from the layer. DNA was detached from the layer using water backwash until the DNA was no longer observed in the eluate.

Product recovery

An 8-column SMB device containing a fluorinated activated carbon solid phase was tested with methanol as solvent to separate ethanol, 2,3-butanediol and acetic acid from the fermentation product and treated as described above. The feed contained 5% ethanol, 1% 2,3-butanediol, 0.8% acetic acid / acetate and the rest were water, medium salts and metals used in the fermentation process. The composition of each stream exiting the SMB apparatus was determined using HPLC technology.

The flow rate of the feed solution was 11 ml / min and the flow rate of the desorbent was 11 ml / min. The step time was 12 minutes and the system was run at a temperature of 75 ° C. The flow rate of the extract stream was optimized to yield the best quality extract, ie the minimum moisture content. 95.7% ethanol and 94.7% 2,3-butanediol from the feed exited the SMB through the extract stream. 41.7% acetic acid / acetate was passed through the extract as acetic acid, with 0.3% of the water from the feed being part of the extract. Three raffinate streams (first raffinate, second raffinate I and second raffinate II) were generated to achieve a stream suitable for recycling with minimal treatment. The flow rates of these raffinate streams were optimized to produce a stream containing minimal metabolites; The optimized flow rates of the first raffinate, secondary raffinate I and secondary raffinate II streams were 5.8 ml / min, 7.5 ml / min and 3.3 ml / min, respectively. The initial raffinate stream contained 4.3% feed ethanol, 5.3% feed 2,3-butanediol and 57.9% feed water. 33.3% of the feed acetic acid / acetate was found in the initial raffienate stream in acetate form. Secondary raffinate I contained 40.7% feed water and the remaining 25% acetic acid / acetate in the form of acetate from the feed. Secondary raffinate II contained 0.1% feed water. 41.7% desorbent methanol was found in the extract stream, 30.7% in secondary raffinate I and 28.5% in second raffinate II.

pH adjustment

Depending on its form, acetic acid / acetate can escape SMB through an extraction or raffinate stream, so it is possible to influence its orientation through pH adjustment of the solution before feeding the solution to SMB. At a pH of about 5, there will be slightly more acetate than acetic acid (pKa for acetic acid is 4.74). Neutralization of the solution is required to ensure that acetic acid exits the SMB as an acetate form; Increasing the pH of the solution to pH 7 or pH 8 will significantly reduce the amount of acetic acid in the solution. Neutralization is achieved through the addition of sodium hydroxide, producing sodium acetate. In order to allow acetic acid to be released in acid form, acidification to a pH close to pH 2 is required, which can be achieved through the addition of acid.

In order to enable the reader to practice the invention without undue experimentation, the invention has been described herein with reference to certain preferred embodiments. Those skilled in the art will recognize that the invention is also capable of interpreting variations and modifications other than those specifically described. It should be understood that the present invention includes all such changes and modifications. In addition, headings, headings, etc. are provided to improve the reader's understanding of the document and should not be read as limiting the scope of the invention. All entire contents of all patent applications, patents and publications cited above and below are incorporated herein by reference.

More particularly, as will be appreciated by those skilled in the art, the implementation of an embodiment of the invention can include one or more additional elements. In various aspects, only those components essential to understanding the present invention may be shown in specific examples or descriptions. However, the scope of the present invention is not limited to the described embodiments, and includes one or more additional steps, and / or one or more substituted steps, and / or systems, and / or methods missing one or more steps. System and / or method.

Reference to any prior art herein should not be intended as any suggestion form or recognition that the prior art forms part of the general knowledge in the field of research in any country around the world.

Throughout this specification and any claims below, the words "comprises", "comprising", and the like, unless otherwise required, are in a generic sense that is contrary to the exclusive meaning, ie "including but not limited to It should be interpreted as ".

Claims (24)

A method for producing and recovering ethanol and 2,3-butanediol from fermentation broth, the method comprising:
a) fermenting a gas base containing CO in a bioreactor containing a culture of one or more carboxydotrophic microorganisms to produce a fermentation broth comprising ethanol and 2,3-butanediol;
b) delivering the fermentation broth to a treatment region, wherein at least a portion of the biomass and soluble protein is removed from the fermentation broth to produce a treated broth stream;
c) delivering at least a portion of the treated broth stream to a simulated moving bed (SMB) module comprising at least one adsorbent;
d) adsorbing at least one of ethanol and 2,3-butanediol onto the adsorbent to yield a raffinate comprising non-adsorbed components of the treated broth stream; And
e) desorbing ethanol and 2,3-butanediol from the adsorbent using at least one desorbent to yield a product stream. The method of claim 1, wherein the treatment region comprises a heat treatment region. The method of claim 2, wherein the treatment region further comprises a filtration region. The method of claim 1, wherein the adsorbent is selected from the group consisting of fluorinated carbon, activated carbon, and modified C18 silica gel. The method of claim 1, wherein the desorbent is selected from the group consisting of ethanol, methanol, propanol, and methyl t-butyl ether. The method of claim 1, wherein the product stream contains water at a concentration of less than 5% by volume. The method of claim 1, wherein the raffinate contains ethanol at a concentration of less than 5% by volume. The method according to claim 1, wherein the microorganism is Moorella ( Morella ), Clostridia , Ruminococcus , Acetobacterium , Eubacterium , Butyribacterium , A method selected from the genus consisting of Oxobacter , Methanosarcina , Desulfotomaculum, and mixtures thereof. The method of claim 1, wherein at least a portion of the raffinate stream is transferred back to the bioreactor. The method of claim 9, wherein one or more nutrients and / or trace elements are added to the raffinate stream prior to delivery to the bioreactor. delete delete delete delete delete delete delete delete delete delete delete delete delete delete

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